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MICB 308 > Koch's Postulates > Flashcards

Koch's Postulates Flashcards

1
Q

What were the theories behind factors that cause disease in the 1800s? (2)

A

-miasma theory: inhaled vapors cause disease
- germ theory: microorganisms cause disease

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2
Q

Koch’s Postulate: First Postulate

A
  • the microorganism must be found in abundance in all organisms suffering from the disease, but should not be found in healthy organisms
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3
Q

Koch’s Postulates: Second Postulate

A
  • the microorganism must be isolated from a diseased organism and grown in pure culture
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4
Q

Koch’s Postulate: Third Postulate

A
  • the cultured microorganism should cause disease when introduced into a healthy host
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5
Q

Koch’s Postulates: Fourth Postulate

A
  • microorganism must be re-isolated from the inoculated diseased experimental host and identified as being identical to the specific causative agent
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6
Q

Challenges to Koch’s Postulates: First (3)

A
  • host susceptibility varies in human and animal populations due to a variety of factors
  • opportunistic bacteria can be found in healthy hosts too
  • may be difficult to isolate bacteria from certain areas, such as brain and bones
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7
Q

Challenges to Koch’s Postulates: Second Postulate (2)

A
  • not all bacteria can be cultured in pure form
  • many bacteria undergo genetic and phenotypic changes in culture environments
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8
Q

Challenges to Koch’s Postulates: Third Postulate (2)

A
  • not all bacterial species are equally virulent
  • closely related bacterial strains can be very different
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9
Q

Challenges to Koch’s Postulates: Fourth Postulate

A
  • re-inoculation of a host may not be possible or ethical
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10
Q

Example challenge to Koch’s Postulates: First Postulate (2)

A
  • Mycobacterium tuberculosis
  • H. pylori
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11
Q

Example challenge to Koch’s Postulates: First Postulate and Mycobacterium tuberculosis (2)

A
  • could not be visualized as it did not gram stain due to differences in cell wall structure
  • new cultivation and staining methods had to be developed
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12
Q

Example challenge to Koch’s Postulates: First Postulate and H. pylori (2)

A
  • causes gastric ulcers, but not all of the ulcers contain H. pylori
  • not all hosts develop disease even if they are carriers
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13
Q

Example challenge to Koch’s Postulates: Second Postulate

A
  • syphilis bacterium
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14
Q

Example challenge to Koch’s Postulates: Second Postulate and Syphilis bacterium (2)

A
  • bacterium has never been cultivated as a pure culture in lab medium
  • rabbit testicles are used as a growth chamber
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15
Q

Example challenge to Koch’s Postulates: Third Postulate (2)

A
  • Helicobacter
  • Salmonella
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16
Q

Example challenge to Koch’s Postulates: Third Postulate and Helicobacter (2)

A
  • Helicobacter mustelae causes disease in animal models, while Helicobacter pylori causes diseases in humans
  • must determine if it is okay to use different bacterial species to satisfy this postulate
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17
Q

Example challenge to Koch’s Postulates: Third Postulate and Salmonella (2)

A
  • Salmonella enterica serovar Typhimurium causes disease in mice while Salmonella enterica serovar Typhi causes disease in humans
  • must determine if it is okay to use different bacterial species to satisfy this postulate
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18
Q

Example challenge to Koch’s Postulates: Fourth Postulate

A
  • is the bacterium directly responsible for the host damage
  • can the bacterium be isolated directly from the site of host damage?
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19
Q

Modern Approaches to Satisfy Koch’s Postulates (4)

A
  • PCR
  • immunohistochemistry and immunofluorescence
  • ELISA
  • eliminate the pathogen and prevent/cure the disease
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20
Q

Modern Approaches to Satisfy Koch’s Postulates: PCR

A
  • PCR and 16S sequencing can be used to amplify and detect bacteria during culturing
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21
Q

Modern Approaches to Satisfy Koch’s Postulates: PCR/16S Sequencing steps (2)

A
  • 16S RNA primers are designed to detect conserved regions, with unique regions in-between that act as barcodes to identify the bacteria
  • PCR is done to amplify the bacterial nucleic acid for detection
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22
Q

Modern Approaches to Satisfy Koch’s Postulates: Immunohistochemistry (2)

A
  • microscopic images of immunohistochemical staining can be used to visualize the pathogen in the tissue directly
  • uses of antibodies against the bacteria with attached reporter enzymes
23
Q

Modern Approaches to Satisfy Koch’s Postulates: Immunofluorescence (2)

A
  • micrographs of cells with fluorescently labelled antibodies against cell markers and bacterial
  • cells infected with bacteria will exhibit colocalization of fluorescent markers, while healthy cells will not
24
Q

Modern Approaches to Satisfy Koch’s Postulates: ELISA (2)

A
  • use of fluorescently labelled antibodies against bacteria to detect pathogens in infection site without culturing
  • Western Blot/SDS-PAGE can be used subsequently
25
Q

ELISA methods (3)

A
  • direct ELISA
  • indirect ELISA
  • capture/sandwich ELISA
26
Q

Modern Approaches to Satisfy Koch’s Postulates: Eliminate the Pathogen and Prevent/Cure the Disease; Why didn’t Koch try this?

A
  • antibiotics did not exist at the time, so treatment was a lot more complicated and time-consuming
27
Q

why is T. pallidum (syphilis bacterium) hard to study using Koch’s Postulates (3)

A
  • bacteria has never been cultivated in the lab
  • does not cause human-like disease in other animals
  • bacterium has several disease stages, each with different symptoms
28
Q

Syphilis: Stage 1 (2)

A
  • painless lesions
  • can be cured with antibiotics
29
Q

Syphilis: Stage 2 (4)

A
  • skin rash on hands and feet
  • spreads across body
  • lasts 2-6 weeks
  • clears on its own without antibiotics
30
Q

Syphilis: Stage 3 (3)

A
  • gumma (large granuloma) forms
  • CNS, eye and heart problems
  • can be cured with antibiotics
31
Q

What does Koch’s Postulates address?

A
  • who/what is causing the disease
32
Q

How do we find out HOW the pathogen is causing the disease? (2)

A
  • must identify mechanism/strategies employed by the bacterium causing the disease
  • must study mechanistic understanding, identification, and infection models
33
Q

virulence factors

A
  • bacterial product or strategy that contributes to the ability of the bacterium to survive in the host/cause infection
34
Q

what are examples of virulence factors from H. pylori (8)

A
  • flagella
  • urease
  • lipopolysaccharides
  • outer proteins for adhesion
  • effector molecules
  • type IV secretion system
  • secretory enzymes
  • exotoxins
35
Q

How can we identify virulence factors?

A
  • genome maps as virulence factors are encoded by genes
36
Q

Molecular Version of Koch’s Postulates: First Postulate

A
  • gene for virulence should be present in the strain of bacteria that cause disease and absent in avirulent strains
37
Q

Molecular Version of Koch’s Postulates: Second Postulate

A
  • (i) knocking out or disruption the gene should reduce virulence, and (ii) introduction of the cloned gene into an avirulent strain should render the avirulent strain virulent
38
Q

Molecular Version of Koch’s Postulates: Third Postulate

A
  • expression of the gene should be demonstrated in human or a relevant model
39
Q

Molecular Version of Koch’s Postulates: Fourth Postulate

A
  • antibodies or a cell-mediated immune response to a virulence factor should be protective
40
Q

Considerations of Models (3)

A
  • relevant experimental system should be chosen as the model
  • the assay that is used only defines the virulence of that assay
  • if the system is flawed, it cannot be extrapolated to humans or other hosts
41
Q

what should we consider before choosing bacterial strains (5)

A
  • representative strains
  • clinical isolates that can be sequences and be tested with genetic tools
  • prototypical wild type strains
  • maintenance of virulence factors that may be metabolically expensive
  • appropriate mutants
42
Q

how can we encourage the maintenance of strain virulence?

A
  • freezing and taking small amounts to study instead of breeding the bacteria for long periods of time
43
Q

what mutants can be used for a study (3)

A
  • spontaneous mutants
  • randomly generated mutants
  • directed mutants
44
Q

spontaneous mutants

A
  • unstable genetic mutants, capable of converting back
45
Q

direct mutants

A
  • more stable mutants
46
Q

what should we consider when choosing hosts for models (3)

A
  • the infection model should be similar; aerosol infection should use an aerosol model
  • tissue distribution of pathology should be similar in both hosts
  • identify closely related pathogens that infect different species
47
Q

advantages of rodent host models (2)

A
  • variability of experiment is smaller due to inbreeding
  • knockouts and transgenic animals exist or can be made easily
48
Q

disadvantages of rodent host models (4)

A
  • lack of variability
  • mice can have different microbiota depending on source
  • mice have different physiologies, habits, and genetics compared to humans
  • human pathogens may not be able to infect rodent model by same route of infections
49
Q

cell lines (2)

A
  • simple model compared to whole organisms
  • generally use epithelial cells, fibroblasts, or monocytes
50
Q

cell lines: advantages (2)

A
  • simple, more controlled, and cheaper than using whole organisms
  • often immortalized by using tumour cells
51
Q

cell lines: disadvantages (3)

A
  • repeated culturing of cell lines can change their properties
  • genes expressed in an organ may not be expressed in tissue culture
  • cell lines are usually not polarized, while cells in organs are
52
Q

polarization of intestinal cells (2)

A
  • cell receptors are asymmetrical
  • apical side (lumen) and basolateral size (tissue/blood) express differential receptors
53
Q

organoids (2)

A
  • new host model that has great potential as it aims to replicate actual organization of human organs
  • derived from human stem cells
54
Q

frequently used host models (7)

A
  • rodents
  • worms (C. elegans)
  • frogs
  • yeast
  • slime mold
  • bees
  • flies (drosophila)

MICB 308 (22 decks)

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